The primary hydrolytic products of dietary fat are fatty acids (FA) and monoacylglycerol (MG). Despite the very large quantities generated in the intestinal lumen, substantial gaps remain in our understanding of the basic mechanisms of MG and FA assimilation by the absorptive cells of the small intestine. These include a molecular level understanding of the individual functions of two highly expressed enterocyte fatty acid-binding proteins (FABP), intestinal FABP and liver FABP (IFABP and LFABP). In our previous studies using in vitro lipid transport kinetics approaches, we provided novel evidence suggesting that IFABP and LFABP are likely to have at least some unique functions in the enterocyte. In the parent proposal, we are using an integrated approach of cellular and animal studies to address the functions of IFABP and LFABP. In Aims 1 and 2, studies are determining the functions and structure/function relationships for IFABP and LFABP in enterocyte transport and metabolism of FA and MG. In our current studies, direct protein transfer and DNA transfection approaches are being used to introduce wild-type and specific mutant forms of the FABPs into cultured cell models that recapitulate the enterocyte phenotype, examining effects on lipid uptake, metabolism, and secretion. In other studies comparing, for the first time, mice null for IFABP or LFABP, we have observed several interesting intestinal phenotypic changes in chow-fed mice, including increased TG synthesis in the IFABP-/- mouse and decreased FA -oxidation in the LFABP knockout. Strikingly, when the animals are fed high saturated fat diets, dramatic whole-body phenotypic differences are observed, with LFABP null mice exhibiting marked obesity and IFABP null mice displaying a completely opposite phenotype--protection against diet-induced obesity! These integrated studies, using a combination of cellular and animal studies, proteomic, lipidomic, and structure-function approaches, are providing entirely novel information about intestinal lipid assimilation, and on the important role of intestinal lipid transport and metabolism in regulating whole-body energy homeostasis. Another important gap in our understanding of intestinal lipid assimilation concerns the metabolic fate of FA and MG in the enterocyte. The absorptive epithelial cell exhibits marked differences in the metabolism of FA added at the dietary or apical (AP) surface of the cell, compared to the basolateral (BL) surface of the cell, and we recently demonstrated striking metabolic polarity for MG metabolism as well. The mechanisms that underlie this compartmentation remain essentially unknown. In our current studies in Aim 3, we are using an integrated approach of cellular and animal studies to determine the mechanisms underlying the metabolic compartmentation of FA and MG in the enterocyte. The overall goal of our research is to provide a molecular level picture of intestinal lipid traffic in order to enable the control of the rate and extent of dietary lipid assimilation and postprandial lipid levels, by modulating specific transport and metabolic processes.